Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) are closely related human pathogens that generate attaching and effacing (AE) lesions to colonize the intestine, damage the epithelium, and promote diarrheal illnesses (Donnenberg and Whittam, J. Clin. Invest. 107, 539-48 (2001); Nataro and Kaper, Clin. Microbiol. Rev. 11, 142-201 (1998)). EPEC and EHEC utilize a type-III-secretion-system to translocate effector proteins into mammalian host cells and form filamentous-actin “pedestals” that characterize AE lesions (Campellone and Leong, Curr. Opin. Microbiol. 6:82-90 (2003); Celli et al., Cell Microbiol. 2:1-9 (2000); Frankel et al., Mol. Microbiol. 30:911-21. (1998)).
A translocated effector protein expressed by both EHEC and EPEC is the translocated intimin receptor, Tir. Tir is delivered into the mammalian plasma membrane where it adopts a hairpin loop conformation (de Grado et al., Cell. Microbiol. 1:7-17 (1999); Hartland et al., Mol. Microbiol. 32:151-158 (1999); Kenny, Mol Microbiol. 31, 1229-41 (1999)) and serves as a receptor for the bacterial surface adhesin intimin (Deibel et al., Mol. Microbiol. 28:463-74 (1998); Kenny et al., Cell 91:511-20 (1997)). The binding of intimin to the central extracellular domain of Tir promotes clustering of the N- and C-terminal cytoplasmic regions and initiates localized actin assembly beneath the plasma membrane (Campellone et al., J. Cell Biol. 164:407-16 (2004)). EHEC Tir and EPEC Tir are approximately 58% identical, and both are critical for the formation of actin pedestals by each pathogen (DeVinney et al., Infect. Immun. 67:2389-98 (1999); Kenny et al., Cell 91:511-20 (1997)).
EPEC Tir is both necessary and sufficient to recruit host Nck SH2/SH3 adaptors via a phosphorylated tyrosine. Nck, in turn, recruits and activates neuronal Wiskott-Aldrich syndrome protein (N-WASP), a key regulator of the Arp2/3 actin-nucleating machinery (Rohatgi et al., J. Biol. Chem. 276:26448-26452 (2001)) required for actin pedestal formation (Lommel et al., EMBO Rep. 2:850-7 (2001)).
EHEC Tir recruits N-WASP and requires N-WASP for efficient pedestal formation (Goosney et al., Infect and Immun. 69: 3315-3322 (2001); Lommel et al., Cell. Microbiol. 6:243-54(2004)). EHEC Tir, however, apparently lacks a phosphotyrosine residue capable of recruiting Nck. Thus, EHEC activates N-WASP and promotes actin pedestal formation using a Nck-independent mechanism. Furthermore, EHEC's Nck-independent mechanism of pedestal formation requires bacterial effectors in addition to Tir.